title: Cecal microbiota transfer rescues the antibiotic-induced acceleration of type 1 diabetes and alteration of intestinal gene expression I

first: Xue-Song

last: Zhang

summary: Early-life antibiotic exposure perturbs the intestinal microbiota, alters innate intestinal immunity, and accelerates type 1 diabetes development in the NOD mouse model. Here we found that maternal cecal microbiota transfer (CMT) to NOD mice with early-life antibiotic perturbation partially rescued the induced T1D acceleration. The restoration effects on the intestinal microbiome were substantial and persistent, remediating the antibiotic-depleted diversity, relative abundance of particular taxa, and metabolic pathways. CMT also protected against perturbed cecal and serum metabolites and normalized innate and adaptive immune effectors. CMT restored patterns of ileal microRNA and histone regulation of gene expression and exon-splicing. Based on the analyses of experimental data, we propose an innate intestinal immune network involving CD44, TLR2, and Reg3g, as well as their multiple microRNA and epigenetic regulators that sense intestinal signaling by the gut microbiota. This regulation affects downstream immunological tone, leading to protection against the tissue-specific T1D injury.

overall_design: Nanostring gene expression analysis was performed on RNA extracted from intestinal epithelial cells of male NOD mice in 3 treatments (control, antibiotic, cecal microbiota transfer) at age of P23

first: Martin

last: Blaser


title: 2-cell small RNA sequencing after paternal dexamethasone injection

status: Public on Nov 25 2020

city: Cambridge

first: Eric

last: Miska

state: Cambridgeshire

address: Gurdon Institute, Tennis Court Rd

department: Cambridge University

country: United Kingdom

institute: Cambridge University

type: Non-coding RNA profiling by high throughput sequencing

summary: 2-cell embryo RNA was sequenced following IVF of naive untreated oocytes with sperm harvested 2 weeks after 2mg/kg dexamethasone or vehicle intraperitoneal injection in males.

overall_design: Examination of sperm small RNA payload in 2-cell embryos to examine effect of paternal dexamethasone injection.

first: Katharina

last: Gapp

first: Alberto

last: Corcoba

first: Fridolin

last: Gross


title: Sperm RNA sequencing after dexamethasone injection

summary: This SuperSeries is composed of the SubSeries listed below.

overall_design: Refer to individual Series


title: Sperm small RNA sequencing 2 weeks after dexamethasone injection

summary: Sperm RNA was sequenced 14 days following 2mg/kg dexamethasone or vehicle intraperitoneal injection.

overall_design: Examination of sperm small RNA payload in pure mature sperm cells, collected from cauda epididymis. These cells were residing in the testis during injection and have passed from Caput to cauda epididymis between the time the animals have been injected and the timepoint of collection.


title: RNA sequencing of nas-38(ok4307), nas-38(tm2655) and wild type C. elegans during L4 lethargus using NextSeq500

status: Public on Nov 19 2020

city: Marburg

first: Marina

middle: Patricia

last: Sinner

state: Hessen

address: Karl-von-Frisch-Str. 8

department: Biology

country: Germany

institute: University of Marburg

type: Expression profiling by high throughput sequencing

summary: The goal of this study was to investigate the differential gene expression between wild types and different nas-38 mutants during lethargus. Extremely prolonged lethargus in nas-38(ok3407) mutants and slightly shortened lethargus in nas-38(tm2655) was observed before.

overall_design: Per condition, three biological replicates were prepared. RNA isolation, library preparation, sequencing and fold change analysis was done by IMGM Laboratories GmbH.

first: Florentin

last: Masurat

first: Henrik

last: Bringmann


title: Single-cell transcriptome conservation in a comparative analysis of fresh and cryopreserved human skin tissue: pilot in localized scleroderma

status: Public on Nov 16 2020

city: Pittsburgh

first: Kathryn

last: Torok

state: Pennsylvania

address: 530 45th St, Rangos 9161 Bay1

department: Peds Rheumatology

country: USA

institute: University of Pittsburgh

summary: In this study, we compare paired fresh (RPMI) compared to frozen (Cryostor®CS10) frozen skin biopsies evaluated by 10X Genomics single cell RNA sequencing.  Comparisons of the cell viability and yield, and transcript expression and yield between the two methodologies were proven to be equivalent.

overall_design: Comparison between fresh and cryopreserved single cell methods in skin

first: Emily

last: Mirizio

first: Tracy

last: Tabib

first: Xinjun

last: Wang

first: Wei

last: Chen

first: Christopher

last: Liu

first: Robert

last: Lafyatis

first: Heidi

last: Jacobe

middle: S


title: bulk RNAseq of SARS-COV2 infected human lung organoid

status: Public on Nov 06 2020

city: los angeles

first: Changfu

last: Yao

state: Ca

address: 8700 Beverly Blvd. AHSP Room A9401

department: Lung and Regenerative Medicine Institutes

institute: cedars-sinai medical center

summary: Coronavirus disease 2019 (COVID-19) is the latest respiratory pandemic resulting from zoonotic transmission of severe acute respiratory syndrome-related coronavirus 2 (SARS-CoV-2). Severe symptoms include viral pneumonia secondary to infection and inflammation of the lower respiratory tract, in some cases causing death. We developed primary human lung epithelial 5 infection models to understand responses of proximal and distal lung epithelium to SARS-CoV-2 infection. Differentiated air-liquid interface cultures of proximal airway epithelium and 3D organoid cultures of alveolar epithelium were readily infected by SARS-CoV-2 leading to an epithelial cell-autonomous proinflammatory response. We validated the efficacy of selected candidate COVID-19 drugs confirming that Remdesivir strongly suppressed viral 10 infection/replication. We provide a relevant platform for studying COVID-19 pathobiology and for rapid drug screening against SARS-CoV-2 and future emergent respiratory pathogens.

sample_organism: Severe acute respiratory syndrome coronavirus 2

overall_design: Organoid generated from primary lung AT2 cells were either transfected with SARS-CoV2 virus or mock were isolated for mRNAseq

first: Apoorva

last: Mulay

first: Barry

middle: R

last: Stripp


title: Modeling Cellular Crosstalk and Organotypic Vasculature Development with Human iPSC-Derived Endothelial Cells (iPS-EC) and Cardiomyocytes (iPS-CM) [scRNA-seq]

city: HELSINKI

first: Emmi

last: Helle

address: Haartmaninkatu 8

country: Finland

institute: University of Helsinki

summary: The aim was to study the effect of 1) coculture of iPS-ECs and iPS-CMs on each cell type and 2) shear stress on iPS-derived endothelial cells (iPS-EC).  The transcriptional landscapes of each cell type were compared.

overall_design: scRNA sequencing analysis of iPS-CMs and iPS-ECs grown in monoculture vs. coculture, and iPS-ECs grown in flow conditions. Two cell lines were used for coculture (HEL47_2 ans HEL24_3), and one for flow (HEL47)

first: Riikka

last: Kivelä

first: Minna

last: Ampuja


title: A human cell atlas of fetal gene expression

status: Public on Nov 01 2020

city: Seattle

first: Junyue

last: Cao

state: WA

address: Foege Building S-210, 3720 15th Ave NE

department: Department of Genome Sciences

institute: University of Washington

summary: The gene expression program underlying the specification of human cell types is of fundamental interest. We generated human cell atlases of gene expression and chromatin accessibility in fetal tissues. For gene expression, we applied three-level combinatorial indexing to >110 samples representing 15 organs, profiling ~4 million single cells. We leveraged the literature and other atlases to identify and annotate hundreds of cell types and subtypes, both within and across tissues. Our analyses focus on organ-specific specializations of broadly distributed cell types (e.g. blood, endothelial, epithelial), sites of fetal erythropoiesis (which surprisingly included the adrenal gland) and integration with mouse developmental atlases (e.g. conserved specification of blood cells). These data comprise a rich resource for the exploration of in vivo human gene expression in diverse tissues and cell types.

overall_design:
single cell RNA-seq profiling of human fetal tissues

***RNA-Seq raw data is to be made available through dbGaP (controlled access) due to privacy concerns:
https://www.ncbi.nlm.nih.gov/gap/?term=phs002003.v1.p1

first: Diana

last: O'Day

first: Hannah

middle: A

last: Pliner

first: Paul

last: Kingsley

first: Mei

last: Deng

first: Riza

middle: M

last: Daza

first: Michael

last: Zager

first: Aldinger

last: Kimberly

first: Ronnie

last: Blecher

first: Fan

last: Zhang

first: Malte

last: Spielmann

first: James

last: Palis

first: Dan

last: Doherty

first: Frank

middle: J

last: Steemers

first: Ian

last: Glass

first: Cole

last: Trapnell

first: Jay

last: Shendure


title: Sperm small RNA sequencing 7 days or 3 hours after dexamethasone injection

first: Eric

last: Miska

summary: Sperm RNA was sequenced 7 days or 3 hours following 2mg/kg dexamethasone or vehicle intraperitoneal injection.

overall_design: Examination of sperm small RNA payload in pure mature sperm cells, collected from cauda epididymis. These cells have passed from Caput to cauda epididymis between the time the animals have been injected and the timepoint of collection.

first: Katharina

last: Gapp

first: Alberto

last: Corcoba


title: Comparative bulk RNAseq analysis of label-retaining chondrocytes (LRCs) of the growth plate versus their progeny

status: Public on Oct 29 2020

city: Houston

first: Noriaki

last: Ono

state: Texas

department: School of Dentistry

institute: University of Texas Health Science Center at Houston

summary: We revealed the unique molecular signature of slow-cycling chondrocytes of the postnatal growth plate.

overall_design: H2B-GFP(high)tdTomato+ label retaining chondrocytes (LRCs) and H2B-GFP(mid-low)tdTomato+ cells were FACS-isolated from postnatal day 49 (P49) growth plates of Col2a1-tTA; TRE-H2B-GFP; Col2a1-creER; R26R-tdTomato quadruple homozygous mice, with Doxycycline from P21 to P49, and two doses of tamoxifen at 3 and 2 days before analyses. mRNA were amplified by SMARTer mRNA-seq kit (Clonetech/Takara) before deep-sequenced by Illumina HiSeq4000.


title: Volatile organic compounds from Bacillus, Serratia and Pseudomonas promote growth and alter the transcriptional landscape of Solanum tuberosum in a passively-ventilated growth system

city: Frankfurt am Main

first: Lukas

last: Jost

address: Altenhöferallee 3

institute: GenXPro GmbH

summary: The interaction of an array of volatile organic compounds (VOCs) termed bacterial volatile compounds (BVCs) with plants is now a major area of study under the umbrella of plant-microbe interactions. Many growth systems have been developed to mediate the investigation of these interactions in vitro. However, each of these systems have their benefits and drawbacks with respect to one another and can greatly influence the end-point interpretation of the BVC effect on plant physiology. To address the need for novel growth systems in BVC-Plant interactions our study investigated the use of a passively-ventilated growth system, made possible via Microbox® growth chambers, to determine the effect of BVCs emitted by six bacterial isolates from the genera Bacillus, Serratia and Pseudomonas. Solid-phase microextraction/GC-MS was utilized to determine the BVC profile of each bacterial isolate when cultured in three different growth media each with varying carbon content. 70 BVCs were identified in total with alcohols and alkanes being the most abundant. When cultured in tryptic soy broth, all six isolates were capable of producing 2,5-dimethylpyrazine, however BVC emission associated with this media were deemed to have negative effects on plant growth. The two remaining media types, namely Methyl Red-Voges Proskeur and Murashige and Skoog, were selected for bacterial growth in co-cultivation experiments with Solanum tuberosum cv. ‘Golden Wonder’. The BVC emissions of Bacillus and Serratia isolates cultured on MR-VP induced alterations in the transcriptional landscape of potato across all treatments with 956 significantly differentially expressed genes. Our results indicate that this novel approach to determining BVC-mediated growth effects on plants is a viable alternative and induced differential growth-promotion based on bacterial isolate and the respective media type on which they were cultured. Surprisingly, genes associated with plant defence were often observed to be downregulated in our system whereas genes associated with photosynthesis and plant cell division were upregulated.

overall_design: Exposure of potato to bacterial volatile organic compounds. Study included 4 treatments to plants; absolute controls, controls grown in proximity to axenic MR-VP media, plants exposed to FZB24 Bacillus VOC's, plants exposed to LAC2 Serratia VOC's. Each treatment consisted of three replicates. Each replicate comprised RNA from 4 individual plants. Plants were exposed to treatments for 4 weeks. Experiments took place in microbox growth chambers, with 18/6 hour day/night exposure to light.

first: Darren

last: Heenan-Daly

first: Simone

last: Coughlan

first: Eileen

last: Dillane

first: Barbara

last: Doyle Prestwich


title: Melanoma mutations are caused by deamination of cytosine in pyrimidine dimers

status: Public on Oct 22 2020

city: Grand Rapids

first: Gerd

middle: P.

last: Pfeifer

state: Michigan

address: 333 Bostwick Ave. NE.

department: Center for Epigenetics

institute: Van Andel Research Institute

summary: The majority of sunlight-associated melanomas carry a unique UV-related C to T mutation signature at dipyrimidine sites. UVB radiation is known to induce cyclobutane pyrimidine dimers (CPDs) as the major form of DNA damage, but the mechanism of how these DNA lesions lead to mutations is unknown. To map the genome-wide distribution of CPDs at single base resolution, we developed a new method, circle damage sequencing (circle-damage-seq). We found that in human cells CPDs form in a specific tetranucleotide sequence context (5’PyPy<>PyT/A), but this alone does not explain the mutation patterns. To test whether mutations arise at CPDs by cytosine deamination, we next applied a modification of circle-damage-seq and mapped UVB-induced cytosine-deaminated CPDs. We observed that the transcription start sites (TSS) are the sequences most protected from CPDs and deaminated CPDs. The strongest spike of CPDs and deaminated CPD lesions was found immediately upstream of the TSS, suggesting a mutation-promoting role of bound transcription factors. Most significantly, the genome-wide dinucleotide and trinucleotide sequence specificity of deaminated CPDs matched the prominent mutation signature found in melanomas. Our data identifies the cytosine-deaminated CPD as the leading premutagenic lesion responsible for the vast majority of mutations in sun-exposure-linked melanomas.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: DNA adducts and deaminations were mapped genome-wide in MEF and HDF cells treated with a dose of 1000 J/m2 and 4000 J/m2 UVB.

first: Seung-Gi

last: Jin

first: Dean

last: Pettinga

middle: P


title: Immune suppressive landscape in a human esophageal squamous cell carcinoma microenvironment

status: Public on Oct 19 2020

city: Shanghai

first: zheyi

last: chen

address: 1665 Kong Jiang Road

country: China

institute: Shanghai Jiao Tong University School of Medicine

summary: Properly applying current immunotherapies or developing novel therapeutic strategies relies on a comprehensive understanding of the immune elements of the tumor microenvironment (TME). In this study, we aim to map at single-cell resolution the immune landscape of esophagus squamous cell carcinoma (ESCC), a common, aggressive, but understudied cancer of the gastrointestinal tract. We obtained a detailed immune cell atlas of the ESCC-associated transcriptome from a total of 80,787 tumor-infiltrating immune cells. Significant intertumoral heterogeneity was found among individual ESSC tumors. Exhausted T cells, exhausted NK cells, regulatory T cells, alternatively activated macrophages, and tolerogenic dendritic cells were discovered as the dominant portions of the immune cells in the TME. Dysfunctional T cells were the major proliferative cell compartment among tumor-infiltrating immune cells. In addition, strong potential interactions of immune cells in tumors may contribute to the immunosuppressive status and the progress of tumor development; a six-gene signature, comprising IFNGR1, IFNGR2, IDO1, IL10, CD163, and CD274, were associated with ESCC survival. Our results provided a comprehensive characterization of tumor-infiltrating immune cells in ESCC, indicating that immunosuppression of various kinds was synchronously responsible for the failure of immunosurveillance in ESCC. Specifically targeting the immunosuppressive pathways could reverse the immune-suppressive status and reactivate anti-tumor immune responses in ESCC.

overall_design: High-dimensional single-cell RNA sequencing (10X Genomics) was used to profile the transcriptomes of total immune cells isolated from seven surgically removed ESSC tumors and their matched adjacent normal tissues. Coupled T cell receptor (TCR) sequencing was also conducted to retrieve information on T cell clonality.

first: Yingxia

last: Zheng

first: Zheyi

last: Chen


title: Transcriptome of Salmonella within Acanthamoeba

status: Public on May 01 2021

city: Orenburg

first: Alexander

last: Balkin

address: 11 Pionerskaya st.

country: Russia

institute: Institute for Cellular and Intracellular Symbiosis

summary: Here, we report global gene expression of S. enterica serovar Typhimurium within Acanthamoeba castellanii by Cappable-Seq

sample_organism: Salmonella enterica subsp. enterica serovar Typhimurium str. ATCC 14028

overall_design: Comparison of control free-living Salmonella with Salmonella within A. castellanii


title: Whole blood transcriptomes of patients receiving digital subtraction angiography

city: BUFFALO

first: Kerry

middle: E

last: Poppenberg

state: NY

address: 875 Ellicott St

institute: University at Buffalo

summary: Circulating whole blood transcriptomes can detect the presence of unruptured cerebral aneurysms. Pending additional testing in larger cohorts, this could serve as a foundation to develop a simple blood-based test to facilitate screening and early detection of cerebral aneurysms.

overall_design: We subjected RNA from peripheral whole blood of 47 patients undergoing cerebral digital subtraction angiogrpahy to next-generation RNA sequencing.

first: Vincent

last: Tutino


title: Derivation of airway basal stem cells from human pluripotent stem cells

status: Public on Oct 14 2020

city: Boston

first: Carlos

last: Villacorta-Martin

state: MA

address: 670 Albany St

institute: Center for Regenerative Medicine at Boston University

summary: The derivation of self-renewing tissue-specific stem cells from human induced pluripotent stem cells (iPSCs) would shorten the time needed to engineer mature cell types in vitro and would have broad reaching implications for the field of regenerative medicine. Here we report the directed differentiation of human iPSCs into putative airway basal cells (“iBCs”), a population resembling the epithelial stem cell of lung airways. Using a dual fluorescent reporter system (NKX2-1GFP;TP63tdTomato) we track and purify these cells over time, as they first emerge from iPSC-derived foregut endoderm as developmentally immature NKX2-1GFP+ lung progenitors which then augment a TP63 program during subsequent proximal airway epithelial patterning.  These cells clonally proliferate, initially as NKX2-1GFP+/ TP63tdTomato+ immature airway progenitors that lack expression of the adult basal cell surface marker, NGFR. However, in response to primary basal cell media, NKX2-1GFP+/ TP63tdTomato+ cells upregulate NGFR and display the molecular and functional phenotype of airway basal stem cells, including the capacity to clonally self-renew or undergo multilineage ciliated and secretory epithelial differentiation in air-liquid interface cultures. iBCs and their differentiated progeny recapitulate several fundamental physiologic features of normal primary airway epithelial cells and model perturbations that characterize acquired and genetic airway diseases. In an asthma model of mucus metaplasia, the inflammatory cytokine IL-13 induced an increase in MUC5AC+ cells similar to primary cells. CFTR-dependent chloride flux in airway epithelium generated from cystic fibrosis iBCs or their syngeneic CFTR-corrected controls exhibited a pattern consistent with the flux measured in primary diseased and normal human airway epithelium, respectively. Finally, multiciliated cells generated from an individual with primary ciliary dyskinesia recapitulated the ciliary beat and ultrastructural defects observed in the donor. Thus, we demonstrate the successful de novo generation of a tissue-resident stem cell-like population in vitro from iPSCs, an approach which should facilitate disease modeling and future regenerative therapies for a variety of diseases affecting the lung airways.Single-cell RNA-Sequencing profiling of human iPSC-derived basal cells, airway epithelium compared to primary human basal cells and airway epithelium.

overall_design: We analyzed (1) iPSC-derived immature airway progenitors, (2) iPSC-derived BCs (iBCs) (3) iBC-derived air-liquid interface(ALI) cultures, (4) primary human basal cells, and (5) primary human basal cells differentiationed in ALI culture. For (1) iPSC-derived immature airway progenitors we differentiated a dual fluorescence iPSC reporter line (NKX2-1GFP;TP63tdTomato), hereaftr BU3 NGPT, in our previously publisehd airway differeniation protocol in imedia containing FGF2, FGF10, dexamethasone, cyclic AMP, 3-isobutyl-1-methylxanthine and Y-27632, hereafter “FGF2+10+DCI+Y”  (McCauley et al., Cell Stem Cell, 2017) (2) iBCs were genereated from the same BU3 NGPT line but cultured in Pneumacult Ex-Plus media supplemented with SMAD inhibitors and Y-27632. Both samples were analyzed by sc-RNA-Seq on day 46 of differentiation. iBCs (GFP+/tdTomato+/NGFR+)  from the BU3 NGPT iPSC line were sorted and plated in ALI conditions to form a pseudostratified airway epithelium on Transwell filters. Freshly isolated, P(0), primary human epithelial cells (basal cells) were cultured in BEGM media and analyzed by Sc-RNA-Seq but also differentiated in air-liquid interface culture in bronchial epithelial growth medium to form a mucociliary epithelium.

first: Finn

last: Hawkins

last: Villacorta

first: Darrell

last: Kotton


title: Transcriptome Profiling of Rectal and Ileal Tissues in Inflammatory Bowel Disease

status: Public on Oct 08 2020

city: Atlanta

first: Kiera

last: Berger

state: GA

address: 950 Atlantic Drive

department: School of Biological Sciences

institute: Georgia Institute of Technology

summary: We report whole mRNA gene expression and Percent Spliced In (PSI) estimates for rectal (n=59) and ileal (n=60) tissue samples of 33 patients with either Crohn's disease (n=19) or ulcerative colitis (n=14)

overall_design: Tissue biopsies were obtained from the rectum and ileum of IBD patients age 8-20 years. Most (n=23) patients donated duplicate biopsies for a total of 4 samples per individual, and at least one sample per location was obtained for all donors. Transcriptome profiling was performed with RNAseq. 119 samples in total were analyzed after QC.

first: Hari

last: Somineni

first: Jarrod

last: Prince

first: Subra

last: Kugathasan

first: Greg

last: Gibson


title: Effect of Akirin2 knockdown on the mRNA profile of human HL60 cells

status: Public on Oct 02 2020

city: Ciudad Real

first: Sara

last: Artigas Jerónimo

state: Ciudad Real

address: Ronda de Toledo 12

department: Sanidad y Biotecnología (SaBio)

country: Spain

institute: Instituto de Investigación en Recursos Cinegéticos (IREC)

summary: The Akirin (AKR) family are transcription cofactors without catalytic or DNA-binding capability with a conserved function in the regulation of multiple biological processes (BPs) throughout the metazoan. Human akirin2 (AKR2) is involved in various BPs in cell type and stimulus-dependent manner by acting as a link between NF-κB and chromatin remodeling complexes (remodelers) for transcriptional regulation. The regulome (transcription factors/cofactors-target genes interactions) plays an important role in the regulation of these BPs and the application of transcriptomics studies can be critical to better understanding the role of this transcription factors/cofactors as AKR2. In this study, we focused in AKR2 regulome to clarify its function as conserved regulatory cofactor. To assess this, RNA-seq analysis were conducted in wild type (WT) and AKR2-knock down (KO) human Caucasian promyelocytic leukemia HL60 cells. Results have shown a different gene expression profile that allowed us to identify immune response genes as putative AKR2 functional complements providing a better understanding of AKR2 regulome and its possible chromatin remodeling function.

overall_design: Total mRNA profile in WT and AKR2 KO HL60 cells (three biological replicates).

last: Artigas-Jerónimo

first: Pilar

last: Alberdi

first: Margarita

last: Villar

first: José

last: de la Fuente


title: Transcriptomics of heart and muscle tissues of transgenic or mdx mice (DMD disease model) with UAA incorporation systems

status: Public on Oct 03 2020

city: Philadelphia

first: Haoran

last: Zhang

address: 415 Curie Boulevard

institute: University of Pennsylvania

summary: We constructed the trans/mdx mice as a novel DMD disease model by crossing transgenic mice and mdx mice. The transgenic and trans/mdx mice have UAA incorporation system that can readthrough the nonsense mutation in the dmd gene with additional UAA. The whole transcriptomics of heart and muscle tissues of 5 mouse groups (n=2) were used to determine whether the introduced UAA incorporation system affect the normal gene expression of mice, or the safety of the system. Groups include: a) wild-type C57BL/6N mice; b) transgenic mice; c) mdx mice; d) trans/mdx mice without UAA; e) trans/mdx mice with UAA (50 mg NAEK every 2-3 day, treated for 4 weeks). The total numbers of expressed genes were around 11,000 in all 5 mouse groups. The expression levels of major genes (logFPKM>0) in 5 mouse groups were also close to each other. These data indicated that the UAA incorporation system was safe and could serve as a potential therapeutic strategy for DMD disease. The transgenic mice with UAA incorporation system we constructed could also cross with other nonsense mutation mice to build more functional animal models.

overall_design: Five groups were designed: a) wild-type C57BL/6N mice; b) transgenic mice; c) mdx mice; d) trans/mdx mice without UAA; e) trans/mdx mice with UAA (50 mg NAEK every 2-3 day, treated for 4 weeks). Two mice were used in each group, whose heart and muscle tissues were collected. Total RNA was extracted by trizol and sent to high-throughput sequencing.

first: Qing

last: Xia

first: Ningning

last: Shi


title: PolyA RNA sequencing of P7 HMGCS2 KO and WT liver

status: Public on Oct 01 2020

city: Kumamoto-City

first: Yuichiro

last: Arima

address: 1-1-1 Honjo

country: Japan

institute: Kumamoto University

summary: To assess the impact of neonatal insuficient ketogenesis, RNA-seq analysis of P7 livers were performed.

overall_design: Liver tissues of Hmgcs2 KO and WT were provided for the analysis.

first: Kenichi

last: Tsujita

first: Shinjiro

last: Hino

first: Mitsuyoshi

last: Nakao


title: Sexual dimorphism in the meiotic requirement for PRDM9: a mammalian evolutionary safeguard

status: Public on Oct 23 2020

city: Bar Harbor

first: Natalie

last: Powers

state: ME

address: 600 Main Street

department: Research

institute: The Jackson Laboratory

type: Genome binding/occupancy profiling by high throughput sequencing

summary: The locations of mammalian recombination hotspots are determined by PRDM9, a zinc finger histone methyltransferase that locally trimethylates histone H3 at residues K4 and K36. Here we report Prdm9-EP, a glu365pro mutation that severely reduces catalytic activity in vivo. This mutation causes sterility with complete meiotic arrest in homozygous males, while homozygous females are able to produce live offspring in natural matings. These H3K4me3 ChIP-seq data from Prdm9-EP homozygous spermatocytes show the extent to which H3K4 methyltransferase activity is compromised by this mutation, while the DMC1 ChIP-seq data show its effect on meiotic double-strand breaks in spermatocytes. For comparison, we mapped previously reported H3K4me3 ChIP-seq data from wild-type C57BL/6J spermatocytes (GSE52628) to mm10, merging the two biological replicates (SRX381465 and SRX381466) before mapping, and proceeding with processing. We did the same with published DMC1 ChIP-seq data, merging three previously reported DMC1 ChIP-seq biological replicates, isolated from wild-type C57BL/6N males (GSE112110; SRX3825301, SRX3825302, and SRX3825303). Processed data files are presented here.

overall_design: We performed ChIP-seq with antibodies against H3K4me3 and DMC1, using chromatin extracted from C57BL/6J-Prdm9-EP homozygous mutant spermatocytes, and compared it to previously generated ChIP-seq data for H3K4me3 and DMC1 from wild-type C57BL/6J or C57BL/6N spermatocytes, respectively.

first: Tanmoy

last: Bhattacharyya


title: A feedback loop between the androgen receptor and 6-phosphogluoconate dehydrogenase (6PGD) drives prostate cancer growth

city: Bedford Park

first: Luke

last: Selth

state: SA

address: Flinders Drive

department: College of Medicine and Public Health

country: Australia

institute: Flinders University

summary: Alterations to the androgen receptor (AR) signalling axis and cellular metabolism are hallmarks of prostate cancer. This study provides insight into both hallmarks by uncovering a novel link between AR and the pentose phosphate pathway (PPP). Specifically, we identify 6-phosphogluoconate dehydrogenase (6PGD) as an AR-regulated gene that is upregulated in prostate cancer. Knockdown of 6PGD impairs growth and elicits death of prostate cancer cells, at least in part due to increased oxidative stress. We investigated the therapeutic potential of targeting 6PGD using 2 specific inhibitors, physcion and S3. Both compounds exhibited substantial anti-cancer activity in multiple models of prostate cancer, including aggressive models of castration-resistant disease that are resistant to contemporary therapy. Mechanistically, 6PGD inhibits AMPK signalling, resulting in increased activity of the anabolic pathways regulated by ACC1 and mTOR. Importantly, the ability of S3 to inhibit prostate cancer growth was recapitulated in prospectively collected tumours using a patient-derived explant (PDE) system. Interestingly, inhibition of 6PGD decreases the expression and activity of AR in both cell lines and PDEs, revealing a novel positive feedback loop between these factors. Supporting the biological relevance of this feedback loop, co-targeting of AR and 6PGD was more effective than targeting either factor alone. Collectively, this work provides new insight into the dysregulated metabolism of prostate cancer and provides impetus for further investigation of co-targeting AR and the PPP as a novel therapeutic strategy.

overall_design: RNA-seq following Enzalutamide treatment or AR knockdown in LNCaP cells.

first: L

last: Butler

first: J

middle: L

last: Gillis

first: D

last: Lynn

first: M

last: Moldovan


title: HIF1α enrichment on KSHV genome in normoxia and hypoxia

first: Rajnish

middle: Kumar

last: Singh

address: 202B Johnson Pavilion 3610 Hamilton walk

department: Otorhinolaryngology

summary: Chromatin Immuno-precipitaion sequencing was performed using HIF1 alpha antibodies to investigate its enrichment on Kaposi's sarcoma associated herpesvirus (KSHV) genome in BC3 cells and purified KSHV infected PBMCs under normoxic or hypoxic conditions

overall_design: KSHV positive BC3 cells or PBMCs infected with purified  KSHV were grown in normoxic or hypoxic (1 % oxygen) conditions in cell culture medium. The cells were harvested and chroamtins were fragmented using sonicator. The fragmented chromatin were clreared with agarose bead and incubated with either HIF1 alpha antibodies or IgG control. The eluted chromatin were used for ChIP sequencing to investigate enrichment of HIF1 alpha on KSHV genome. Data analysis was performed using CLC Genomics workbench 12 software.

first: Erle

last: Robertson

middle: K


title: Pathogenesis, miR-122 gene-regulation, and protective immune responses after acute infection of horses with equine hepacivirus

status: Public on Sep 30 2020

city: Copenhagen N

first: Troels

middle: K H

last: Scheel

state: Copenhagen

address: Blegdamsvej 3

department: Institute for Immunology and Microbiology

country: Denmark

institute: University of Copenhagen

summary: Equine hepacivirus (EqHV) is the closest genetic relative of hepatitis C virus (HCV) and shares features of genome organization, hepatotropism, persistent infection, and the ability to cause liver disease. As such, EqHV studies are important both in order to understand equine liver disease, and as an outbred animal model for HCV pathogenesis and immune responses. Here, we characterize the natural history and immune response to EqHV infection. Seven horses were experimentally inoculated with EqHV, monitored for 6 months, and challenge inoculated with the same, and subsequently a divergent EqHV inoculum. Clearance was the primary outcome (6 of 7) and was associated with subclinical hepatitis characterized by lymphocytic infiltrate and individual hepatocyte necrosis. Seroconversion was delayed and antibody titers waned slowly. Resolving horses developed non-sterilizing immunity resulting in short duration of infection upon challenge. Unlike those observed in acutely HCV-infected patients, peripheral blood mononuclear cell responses in horses were minimal, although EqHV specific T-cells were identified. In contrast, an interferon stimulated gene signature was detected in the liver during EqHV infection, which is similar to acute HCV in humans. EqHV, similarly to HCV, is stimulated by direct binding of the liver-specific microRNA, miR-122. Surprisingly, we found that EqHV infection sequesters enough miR-122 to functionally affect gene regulation in the liver. This RNA-based mechanism thus could have consequences for pathology. Conclusion: EqHV infection in horses typically has an acute resolving course, and the immune response attenuates subsequent infections lasting for at least a year. This could have important implications to achieve the first goal of an HCV vaccine; to prevent chronicity while accepting acute resolving infection after challenge.

overall_design: Seven horses were experimentally infected with equine hepacivirus (EqHV). RNA-Seq on PBMC and liver biopsies was performed pre-inoculation (PRE), during early acute infection (ACUTE), near seroconversion (SC), during falling viremia (FV), and at week 26 (POST). Six horses cleared the infection within the 26 weeks, and these were challenged with homologous serum (week 26) and heterologous serum (week 30-32). Additional challenge time points were included for horses that became positive during challenge and where sampling was possible (horses A, P, R for liver, horses A, N, P, R for PBMCs). One replicate was processed per horse per time point. PBMC libraries were poly-A selected; liver biopsy libraries were total RNA-seq (ribo-depletion).

first: Joy

last: Tomlinson

first: Raphael

last: Wolfisberg

first: Ulrik

last: Fahnøe

first: Roosheel

last: Patel

first: Sheetal

last: Trivedi

first: Arvind

last: Kumar

first: Himanshu

last: Sharma

first: Louise

last: Nielsen

first: Sean

last: McDonough

first: Jens

last: Bukh

first: Bud

middle: C

last: Tennant

first: Amit

last: Kapoor

first: Brad

last: Rosenberg

first: Charles

last: Rice

first: Thomas

last: Divers

first: Gerlinde

last: Van de Walle


title: Whole genome bisulfite sequencing of apoptotic and male-differentiated subpopulations of E13.5 wild-type murine male germ cells

city: San Francisco

first: Daniel

last: Nguyen

address: 513 Parnassus Ave

institute: University of California San Francisco

type: Methylation profiling by high throughput sequencing

summary: Heterogeneity within murine prospermatogonia manifests as certain populations either die or survive during a developmentally scheduled apoptotic event at E13.5. We have identified that this apoptosis is clonal, suggesting that apoptotic fate is heritably enforced among populations of germ cells. To determine how subpopulations of germ cells differ, we used single-cell RNA sequencing to identify two diametric states: an apoptotic (AP) versus a male-differentiated (MD) subpopulation that differ primarily by the expression of critical male-differentiation genes known to be epigenetically regulated. We isolated AP and MD germ cells from E13.5 male embryos for whole genome bisulfite sequencing to compare how methylation differs between these populations. We find that AP germ cells are relatively hypermethylated at a whole-genome level and, importantly, are significantly hypermethylated at specific genome-reprogramming responsive (GRR) loci. Because these GRR genes facilitate male differentiation, we conclude that inappropriate demethylation as germ cells epigentically reprogram in the period preceding apoptosis produces an aberrantly male-differentiated clone that is fated for elimination. This quality-control paradigm therefore links fidelitous epigenetic reprogramming to germ cell selection.

overall_design: AP and MD germ cell populations were FACS-isolated from E13.5 male murine embryos from biological replicates. One replicate contained 2000 cells per each population while the second replicate contained 6000 cells per each population. Sequencing was completed for whole-genome analysis as well as specifically for 45 GRR genes and LINE1 elements.

first: Suping

last: Peng

first: Diana

last: Laird


title: Effects of sheared chromatin length on ChIP-seq quality and sensitivity

city: University Park

first: Ross

last: Hardison

state: PA

address: 303 Wartik Lab

institute: Pennsylvania State University

summary: Chromatin immunoprecipitation followed by massively parallel, high throughput sequencing (ChIP-seq) is the method of choice for identifying, on a genome-wide scale, the segments of DNA bound by specific transcription factors (TFs) or in chromatin with particular histone modifications. However, the quality of ChIP-seq datasets vary over a wide range, with a substantial fraction being of intermediate to poor quality. Such experimental variability can lead to many false positives or false negatives, impairing the ability to interpret the data. Thus, it is important to discern and control the factors that contribute to variation in ChIP-seq. In this study we focus on the sonication step to produce sheared chromatin, a variable controllable by the user and applicable to all ChIP-seq protocols. We systematically varied the amount of shearing of fixed chromatin from a mouse erythroid cell line, carefully measured the distribution of resultant fragment lengths using the Agilent Bioanalyzer 2100, and then immunoprecipitated these batches of chromatin using highly specific antibodies against either TAL1 or CTCF. We found that the level of sonication, which was affected by both the number of sonication cycles, as well as the starting cell number, had a pronounced impact on the quality of resulting ChIP-seq signals. Specifically, over-sonication led to degradation of quality (e.g. increased background and reduction in signal), while the impact of under-sonication of chromatin differed between the two transcription factors, leading to the loss of sites occupied by TAL1 but not those bound by CTCF. We leveraged these findings to produce a set of CTCF ChIPs-seq datasets in primary hematopoietic progenitor cells, including several rare cell types. Together, these results suggest that the amount of sonication is a key variable in success of ChIP-seq experiments, and that carefully monitoring the level of chromatin sonication is one way to improve ChIP-seq quality and reproducibility, which in turn facilitates low input ChIP-seq in rare cell types.

overall_design:
We systematically varied the extent of sonication of fixed chromatin from G1E-ER4+E2 cells, carefully measured the distribution of resultant fragment lengths using the Agilent Bioanalyzer 2100, and then immunoprecipitated these batches of chromatin using highly specific antibodies against either TAL1 or CTCF. Following ChIP-seq library preparation, these samples were sequenced on the Illumina NextSeq 500. The datasets were then examined by both subjective visual assessments of the signal tracks at various loci on the genome browser, as well as established, genome-wide, objective quality metrics to determine the effects of sheared chromatin length on ChIP-seq quality and sensitivity. These findings were then leveraged to produce a set of CTCF ChIP-seq datasets in 8 bone marrow-derived, hematopoietic stem and progenitor cells, including several rare cell types.
Samples GSM946520 (GSE36028, GSE49847, GSE51338) and GSM995439 (GSE36029, GSE49847, GSE51338) were used as the input samples for the G1E-ER4+E2 samples.

first: Cheryl

last: Keller

first: Alexander

middle: Q

last: Wixom

first: Elisabeth

middle: F

last: Heuston

first: Belinda

last: Giardine

first: Chris

last: Hsiung

first: Maria

last: Long

first: Amber

last: Miller

first: Stacie

last: Anderson

first: April

last: Cockburn

first: Gerd

last: Blobel

first: David

last: Bodine


title: Transcriptome analysis identifies Doublesex and Mab-3 Related Transcription Factor (DMRT3) in nasal polyp epithelial cells of patients suffering from Nonsteroidal anti-inflammatory drugs-  Exacerbated Respiratory Disease (N-ERD)

status: Public on Sep 22 2020

city: CDMX

first: Luis Manuel

last: Teran Juarez

state: Ciudad de México

address: Calz. de Tlalpan 4502, Belisario Domínguez Secc 16, Tlalpan,

department: immunogenetics and allergy

country: Mexico

institute: Instituto Nacional de Enfermedades Respiratorias Ismael Cosío Villegas

summary:
Objective: To establish the transcriptome of the nasal polyp airway epithelial cells derived from N-ERD patients to uncover the gene expression patterns during this disease.
Methods: Nasal airway epithelial cells were isolated from 12 N-ERD polyps and 9 N-ERD non polyp nasal mucosa as controls from the same subjects. RNA was sequenced on the Illumina HiSeq 2000. Potential gene candidate DMRT3 was selected from the differentially expressed genes for validation.
Results: Comparative transcriptome profiling of nasal epithelial cells was achieved in N-ERD. 18 genes had twofold mean regulation expression differences or greater. 5 genes were upregulated including DMRT3 and 12 genes were down-regulated. Differentially regulated genes included inflammation, defense and immunity. Significantly enriched pathways were metabolic process and embryonic development. ELISA results of DMRT3 in N-ERD patients was significantly upregulated when compared to controls (p= 0.03). IHC of N-ERD nasal polyps localised DMRT3 and was predominantly released in the airway epithelia. These results corroborate with our findings.
Conclusion: Findings suggest that DMRT3 could be potentially involved in nasal polyps development in N-ERD patients. Known functions of DMRT3 include nucleic acid binding and highly expressed during embryonic development. Several genes are downregulated, hinting dedifferentiation phenomenon in N-ERD polyps. However, further studies are required to confirm the exact mechanism of polyps formation in N-ERD patients.

overall_design: mRNA profiles of N-ERD polyps and non polyps N-ERD nasal mucosa.

first: V

last: Priyadharshini

first: Marcos

last: J-Chobillon

first: Jos

last: de Graaf

first: Raúl

last: Gutiérrez de Velasco

first: Christina

last: Gratziou

first: Fernando

last: Jiménez

first: Luis

last: Teran


title: Co-culture with mouse embryonic fibroblasts improves maintenance of metabolic function of human small hepatocyte progenitor cells

status: Public on Sep 11 2020

city: Madison

first: Srikumar

last: Sengupta

state: WI

address: 330 N Orchard St

department: Regenerative Biology

institute: Morgridge Institute for Research

summary: Small hepatocyte progenitor cells (SHPCs) can be derived from primary adult human hepatocytes on mouse embryonic fibroblasts (MEFs). SHPCs derived and cultured on MEFs retain their differentiated morphology and gene expression profiles over several passages. SHPCs in MEF co-culture metabolize acetaminophen to all of its major metabolites at rates equal to mature adult primary human hepatocytes.

overall_design: Examination of SHPC co-culture on MEFs for better retention of metabolic functions

first: Brian

last: Johnson

first: Morten

last: Seirup

first: Hamisha

last: Ardalani

first: Bret

last: Duffin

first: Gregory

last: Barrett-Wilt

first: Jennier

last: Bolin

first: John

last: Steill

first: Scott

last: Swanson

first: Amy

last: Freitag

first: Cara

last: Argus


title: Cytoplasmic mRNA and Heavy polysome-associated mRNAs from U251 Cells treated with Control or eIF3e siRNAs

status: Public on Sep 09 2020

city: Trento

first: Erik

last: Dassi

state: TN

address: Via Sommarive, 9

department: CIBIO

country: Italy

institute: University of Trento

summary: Transcriptional profiling of Cytoplasmic mRNA and Heavy polysome-associated mRNAs extracted from U251 Cells treated with control siRNA or siRNA targeting eIF3e . Goal was to determine the effect of eIF3e knockdown on translation of specific mRNAs.

overall_design: Four-condition experiment, cytoplasmic mRNA and polysome associated-mRNA in U251 cells siScr VS sieIF3e . Biological replicates: 4 replicates

first: Julie

last: Sesen

first: Anne

last: Cammas

first: Nicolas

last: Skuli

first: Stefania

last: Millevoi


title: miRNome changes in the mouse kidney after unilateral ischemia-reperfusion injury and delayed contralateral nephrectomy

status: Public on Sep 01 2020

city: Budapest

first: Pál

last: Tod

state: Budapest

address: Szigony utca 43

country: Hungary

institute: Institute of Experimental Medicine

summary: Comparing the effect of unilateral ischemia-reperfusion injury (IRI) or sham operation (sIRI) with delayed contralateral nephrectomy (Nx) or sham operation (sNx) in mouse kidney. IRI was performed on day 0 and the contralateral kidney was removed on day 7. Mice were sacrificed on day 8. Four animals were selected from the sham IRI-sham Nx and sham IRI-Nx groups and six animals were selected from the IRI-sham Nx and IRI-Nx groups for miRNA microArray analysis on the base of their proinflammatory (TNF-α and IL-6 and CCL2) and immune system-related (Complement component 3) mRNA expression levels.

overall_design: Four-condition design: sham IRI-sham Nx (sIRI-sNx) n=4, sham IRI-Nx (sIRI-Nx) n=4, IRI-sham Nx (IRI-sNx) n=6, IRI-Nx (IRI-Nx) n=6

first: Beáta

last: Róka

first: Tamás

last: Kaucsár

first: Éva

middle: N

last: Bukosza

first: Gábor

last: Szénási

first: Péter

last: Hamar


title: A first-in-class, humanized antibody targeting alternatively spliced tissue factor: preclinical evaluation in an orthotopic model of pancreatic ductal adenocarcinoma

city: Cincinnati

first: Kateryna

last: Matiash

state: Ohio

address: 3125 Eden Ave

department: Hematology/Oncology

institute: University of Cincinnati

summary: RNA sequencing of 9 samples (3 per group) generated 30 million 150 base long paired end reads revealed differential isoform expression of about 1100 gene transcripts. Hg19 was used as a reference genome for alignment, transcriptome index was created in kallisto using Ensembl cDNA sequences for the reference genome. This index was then used to quantify transcript abundance in raw counts and transcript per million (TPM). We report a suppression in the expression of genes associated with cell cycle regulation (most strongly HBGEF, STN1, ECT2, PLCD3, and RHOC).  Suppression of HBGEF and STN1 in hRabMab1 treated tumors was validated by qRT-PCR.

overall_design: Systemic delivery of a humanized monoclonal antibody targeting human alternatively spliced tissue factor (asTF): asTF-targeting hRabMab1 antibody treatment compared to vehicle or hIgG1 controls in nude mice after orthotopic injection of human Pt45.P1 pancreatic ductal adenocarcinoma cells. RNA was isolated from crushed tumors representing the median 3 tumor volumes from each treatment group (Vehicle, hIgG1, hRabMab1).  RNA library preparation, next generation sequencing, and sequence alignment were performed by the CCHMC DNA Sequencing and Genotyping Core, Cincinnati, Ohio.

first: Clayton

last: Lewis

first: Aniruddha

last: Karve

first: Timothy

last: Stone

first: Bruce

last: Aronow

first: Vladimir

middle: Y

last: Bogdanov


title: Delineating mechanisms of immune evasion in lung carcinoma in situ

city: London

first: Vitor

middle: H

last: Teixeira

address: 5 University Street

department: Medicine

institute: University College London

summary: Background: Lung carcinoma-in-situ (CIS) lesions are the pre-invasive precursor to lung squamous cell carcinoma. However, only half progress to invasive cancer in three years, while a third spontaneously regress. Molecular profiling techniques have identified marked differences between progressive and regressive lesions. Here we present laser-captured stromal tissue, adjacent to previously profiled CIS lesions, to examine the role of the tumour microenvironment in spontaneous lesion regression. All samples in this dataset are paired with matched CIS lesions in dataset GSE108082.

overall_design: Progressive and regressive lung CIS lesions were laser-captured and their transcriptome interrogated. This data set represents matched laser-captured stromal tissue adjacent to previously profiled CIS lesions (GSE108082). Differential expression analysis was performed to identify stromal differences between progressive and regressive lesions.

first: Adam

last: Pennycuick


title: Screening of PCRP transcription 1 factor antibodies in biochemical assays

city: Chicago

first: Ali

last: Shilatifard

state: IL

address: 320 E Superior St

department: Department of Biochemistry and Molecular Genetics

institute: Northwestern University Feinberg School of Medicine

summary: Antibodies offer a powerful means to interrogate specific proteins in a complex milieu, and where epitope tagging is impractical. However, antibody availability and reliability are problematic. The Protein Capture Reagents Program (PCRP) generated over a thousand renewable monoclonal antibodies against human-presumptive chromatin proteins in an effort to improve reliability. However, these reagents have not been widely field-tested. We therefore screened their ability in a variety of assays. 887 unique antibodies against 681 unique chromatin proteins, of which 605 are putative sequence-specific transcription factors (TFs), were assayed by ChIP-exo. Subsets were further tested in ChIP-seq, CUT&RUN, STORM super-resolution microscopy, immunoblots, and protein binding microarray (PBM) experiments. At least 6% of the tested antibodies were validated for use in ChIP-based assays by the most stringent of our criteria. An additional 34% produced data suggesting they warranted further testing for clearer validation. We demonstrate and discuss the metrics and limitations to antibody validation in chromatin-based assays.

overall_design: ChIP-sequencing in HCT116 cells for hybridomas generated by NIH's Protein Capture Reagent Program. For NRF1, two hybridomas were tested with and without RNAi. For HSF1, seven hybridomas were tested at both 37 degrees and after 1 hour of heat shock (43°C).

first: Edwin

middle: R

last: Smith


title: High throughput joint profiling of chromatin accessibility and protein levels in single cells

status: Public on Sep 08 2020

city: New York

first: Caleb

last: Lareau

state: New York

address: 417 E 68th St, Zuckerman - ZRC 1132

institute: Memorial Sloan Kettering

summary: This SuperSeries is composed of the SubSeries listed below.

overall_design: Refer to individual Series


title: Enhancer Release and Retargeting” Activates Disease Susceptibility Genes (ChIP-Seq)

status: Public on Aug 31 2020

city: Houston

first: Jiaofang

last: Shao

state: Texas

address: 6431 Fannin St

department: Department of Biochemistry and Molecular Biology

institute: McGovern Medical School, University of Texas Health Science Center

summary: We used genome wide ChIP-seq to examine the chromatin histone changes after the knockdown of one of cohesin subunit, RAD21, in MCF7 breast cancer cells.

overall_design: Examination of h3k4me1,h3k4me3 histone modifications by ChIP-seq.

first: Soohwan

last: Oh

first: Wenbo

last: Li


title: Enhancer Release and Retargeting” Activates Disease Susceptibility Genes


title: Plasmodium falciparum increased time in circulation underlies persistent asymptomatic infection in the dry season

status: Public on Aug 20 2020

city: Hamilton

first: Craig

last: Martens

state: MT

address: 903 S. 4th St

department: DIR

institute: NIAID

summary: The dry season is a major challenge for Plasmodium falciparum parasites in many malaria endemic regions, where water availability limits mosquitoes to only part of the year. How P. falciparum bridges two transmission seasons months apart, without being cleared by the host or compromising host survival is poorly understood. Here we show that low levels of P. falciparum parasites persist in the blood of asymptomatic Malian individuals during the 5- to 6-month dry season, rarely causing symptoms and minimally affecting the host immune response. Parasites isolated during the dry season are transcriptionally distinct from those of subjects with febrile malaria in the transmission season, reflecting longer circulation within each replicative cycle, of parasitized erythrocytes without adhering to the vascular endothelium. Low parasite levels during the dry season are not due to impaired replication, but rather increased efficiency of splenic clearance of longer-circulating infected erythrocytes. We propose that P. falciparum virulence in areas of seasonal malaria transmission is regulated so that the parasite decreases its endothelial binding capacity, allowing increased splenic clearance and enabling several months of subclinical parasite persistence.

overall_design: RNA-Seq of Plasmodium falciparum collected from 12 individuals during dry season compared to Plasmodium falciparum collected from 12 individuals in ensuiing transmission season (wet).

first: Carolina

last: Andrade

first: Hannah

last: Fleckenstein

first: Richard

last: Thomson-Luque

first: Safiatou

last: Doumbo

first: Nathalia

last: Lima

first: Carrie

last: Anderson

first: Julia

last: Hibbert

first: Christine

last: Hopp

first: Tuan

last: Tran

first: Shanping

first: Moussa

last: Niangaly

first: Didier

last: Doumtabe

first: Jeff

last: Skinner

first: Dan

last: Sturdevant

first: Stacy

last: Ricklefs

first: Kimmo

last: Virtaneva

first: Muhammad

last: Asghar

first: Manijeh

middle: V

last: Homann

first: Louise

last: Turner

first: Joana

last: Martins

first: Erik

last: Allman

first: Marie-Esther

last: N'Dri

first: Volker

last: Winkler

first: Manuel

last: Llinás

first: Catherine

last: Lavazec

first: Anna

last: Farnert

first: Kassoum

last: Kayentao

first: Aissata

last: Ongoiba

first: Thomas

last: Lavstsen

first: Nuno

last: Osório

middle: D

last: Otto

first: Mario

last: Recker

first: Boubacar

last: Traore

first: Peter

last: Crompton

first: Silvia

last: Portugal


title: Dynamin-related protein 1 deficiency accelerates lipopolysaccharide-induced acute liver injury and inflammation in mice

status: Public on Aug 28 2020

city: Fukuoka

first: Lixiang

last: Wang

state: Fukuoka

address: 3-1-1, Maidashi, Higashi-ku

department: Department of Medicine and Bioregulatory Science

institute: Graduate School of Medical Sciences, Kyushu University

summary: Mitochondrial fusion and fission, which are strongly related to normal mitochondrial function, are referred to as mitochondrial dynamics. Mitochondrial fusion defects in the liver cause a non-alcoholic steatohepatitis-like phenotype and liver cancer. However, whether mitochondrial fission defect directly impair liver function and stimulate liver disease progression, too, is unclear. Dynamin-related protein 1 (DRP1) is a key factor controlling mitochondrial fission. We hypothesized that DRP1 defects are a causal factor directly involved in liver disease development and stimulate liver disease progression. Drp1 defects directly promoted endoplasmic reticulum (ER) stress, hepatocyte death, and subsequently induced infiltration of inflammatory macrophages. Drp1 deletion increased the expression of numerous genes involved in the immune response and DNA damage in Drp1LiKO mouse primary hepatocytes. We administered lipopolysaccharide (LPS) to liver-specific Drp1-knockout (Drp1LiKO) mice and observed an increased inflammatory cytokine expression in the liver and serum caused by exaggerated ER stress and enhanced inflammasome activation. This study indicates that Drp1 defect-induced mitochondrial dynamics dysfunction directly regulates the fate and function of hepatocytes and enhances LPS-induced acute liver injury in vivo.

overall_design: We have reported that disruption of mitochondrial fission by deletion of Drp1 leads to an alteration in expression of the genes involved in the immune system in the liver and significant elevation of pro-inflammatory cytokines was detected in Drp1 knockout cells. To fully reveal the gene expression altered by Drp1 deletion, we conducted an independent microarray analysis with primary hepatocyte harvested 24 hours after seeding.

first: Masatoshi

last: Nomura


title: Single-cell RNA-seq of the goat ovarian granulosa cells samples from high and low fertility groups

status: Public on Aug 01 2020

city: Qingdao

first: Yong

last: Zhao

state: Shandong

address: 700 Changcheng Road

department: College of Life Sciences

institute: Qingdao Agricultural University

summary: The goal of this study was designe to search for the granulosa cells contribution on fertility. Single-cell RNA-seq of the goat ovarian granulosa cells samples from high fertility group (>three babys per birth) and low fertility group (< two babys per birth). And each group was divided into three subgroup based in the size of follicles: large, medium, and small follicles.

first: Shen

last: Li

first: Lingjiang

last: Min


title: Next Generation Sequencing of LSK EpCAM+ and EpCAM- of Zeb1-/-

city: AlDawadmi

first: Alhomidi

last: Almotiri

address: King Abdulaziz Rd

country: Saudi Arabia

institute: Shaqra University

summary: Zeb1, a zinc finger E-box binding homeobox epithelial-mesenchymal (EMT) transcription factor, confers properties of ‘stemness’, such as self-renewal, in cancer. Yet little is known about the function of Zeb1 in adult stem cells. Here, we used the hematopoietic system, as a well-established paradigm of stem cell biology, to evaluate Zeb1 mediated regulation of adult stem cells. We employed a conditional genetic approach using the Mx1-Cre system to specifically knockout (KO) Zeb1 in adult hematopoietic stem cells (HSCs) and their downstream progeny. Acute genetic deletion of Zeb1 led to rapid onset thymic atrophy and apoptosis driven loss of thymocytes and T cells. A profound cell-autonomous self-renewal defect and multi-lineage differentiation block was observed in Zeb1 KO HSCs. Loss of Zeb1 in HSCs activated transcriptional programs of deregulated HSC maintenance and multi-lineage differentiation genes, and of cell polarity, consisting of cytoskeleton, lipid metabolism/lipid membrane and cell adhesion related genes. Notably, Epithelial cell adhesion molecule (EpCAM) expression was prodigiously upregulated in Zeb1 KO HSCs, which correlated with their enhanced cell survival capacity and diminished differentiation in transplantation. Thus, Zeb1 acts as a crucial transcriptional regulator in hematopoiesis, co-ordinating HSC self-renewal and multi-lineage differentiation fates, in part, via EpCAM repression.

overall_design: EpCAM+ and EpCAM- fractions from Zeb1-/- LSK cells were isolated from 8-to-12-week old mice (4 mice per genotype)

first: Hamed

last: Alzahrani

first: Neil

last: Rodrigues


title: Sperm long RNA sequencing 14 days after dexamethasone injection

status: Public on Jul 15 2020

city: Cambridge

first: Eric

last: Miska

state: Cambridgeshire

address: Gurdon Institute, Tennis Court Rd

department: Cambridge University

institute: Cambridge University

summary:
This study investigates the impact of Dexamethasone on sperm RNA in mice. Using RNA sequencing we find that Dexamethasone impacts the sperm payload significantly 2 weeks post intra peritoneal administration.
Sperm RNA was sequenced 14 days following 2mg/kg dexamethasone or vehicle intraperitoneal injection.

overall_design: Examination of sperm long RNA payload in pure mature sperm cells, collected from cauda epididymis. Cells were in testis at time of injection. These cells have passed from Caput do cauda epididymis between the time the animals have been injected and the timepoint of collection.

first: Katharina

last: Gapp

first: Alberto

last: Corcoba

first: Fridolin

last: Gross


title: Fusion protein EWS-FLI1 is incorporated into a protein granule in cells

status: Public on Jul 23 2020

city: Tucson

first: Jacob

middle: C.

last: Schwartz

state: AZ

address: 1501 N Campbell Ave, LSN649

department: Pharmacology

institute: University of Arizona

summary: Ewing sarcoma is driven by fusion proteins containing a low complexity (LC) domain that is intrinsically disordered and a powerful transcriptional regulator. The most common fusion protein found in Ewing sarcoma, EWS-FLI1, takes its LC domain from the RNA-binding protein EWSR1 (Ewing Sarcoma RNA-binding protein 1) and a DNA-binding domain from the transcription factor FLI1 (Friend Leukemia Virus Integration 1). The LC domain in EWS-FLI1 can bind RNA polymerase II (RNA Pol II) and can self-assemble through a process known as phase separation. The ability of EWSR1 and related RNA-binding proteins to assemble into ribonucleoprotein granules in cells has been intensely studied but the role of phase separation in EWS-FLI1 activity is less understood. We investigated the overlapping functions of EWSR1 and EWS-FLI1 in controlling gene expression and tumorigenic cell growth in Ewing sarcoma, and our results suggested that these proteins function closely together. We then studied the nature of interactions among EWS-FLI1, EWSR1, and RNA Pol II. We observed EWSR1 and RNA Pol II to be present in protein granules in cells. We then identified protein granules in cells associated with the fusion protein, EWS-FLI1. The tyrosine residues in the LC domain are required for the abilities of EWS-FLI1 to bind its partners, EWSR1 and RNA Pol II, and to incorporate into protein granules. These data suggest that interactions among EWS-FLI1, RNA Pol II, and EWSR1 in Ewing sarcoma can occur in the context of a molecular scaffold found within protein granules in the cell.

overall_design: Comparison of transcript levels for knockdown of two proteins or both in Ewing sarcoma.

first: Nasiha

middle: S

last: Ahmed


title: Transcriptome analysis of SFP45 knockdown cells

status: Public on Jul 30 2020

city: Toyoake

first: Rei

last: Yoshimoto

state: Aichi

address: 1-98 Dengakugakubo, Kutsukake

department: Institute for Comprehensive Medical Science

institute: Fujita Health University

summary: Examination of transcriptome of SPF45 knockdown cells by Ribosomal RNA depleted RNA-Seq

first: Kazuhiro

last: Fukumura


title: A Candidate Gene Identified in Converting Platycoside E to Platycodin D by Correlating Main Metabolites Content with Gene Expression Variation between Callus and Leaf of Platycodon grandiflorus

city: Hefei

first: Shihai

last: Xing

state: Anhui

address: No.1 Qianjiang Road, Xinzhan District

institute: Anhui University of Chinese Medicine

summary: Platycodon grandiflorus is a perennial traditional Chinese herb, which has more than 100 compounds including triterpenoid saponins, polysaccharides and phenolic acid etc. In this study, a (L9(34)) orthogonal experiment determined the optimal explants is stem with nodes and the optimum full medium is MS + NAA 1.0 mg/L + 6-BA 0.5 mg/L as best protocol for callus induction of P. grandiflorus.The content of platycodin D, platycoside E and total polysaccharides between callus and plant organs is significantly difference, and the distribution of platycodin D, platycoside E and total polysaccharide in whole plant has been clarified. Meanwhile, the content of platycodin D and total polysaccharide were found higher in calli than in leaves or the whole plant. While, the content of platycoside E in calli is very lower than in leaves. In order to screen candidate genes involved in platycoside E and platycodin D conversion, a RNA-Seq analysis between calli and leaves was performed. Correlating the content of platycodin D and platycoside E between calli and leaves with the gene expression level involving in triterpenoid saponin biosynthesis pathway, four cluster genes with high amino acid similarity were screened as the putative sequences of β-Glucosidase gene for converting platycoside E to platycodin D. It was found that the expression level of genes encoding enzymes involved in saponins and polysaccharides biosynthesis was co-expression with the content of the corresponding metabolite. Besides, some transcription factors predicted to regulate the expression of key enzymes involving in triterpernoid saponins and polysaccharides biosynthesis pathway of P. grandiflorus.

overall_design: Try to find β-glucosidase gene in plants to transform Platycoside E to Platycoside D

first: Xinglong

last: Su


title: Fosl1 regulates mesenchymal GBM plasticity

city: Madrid

first: Massimo

last: Squatrito

address: Calle Melchor Fernandez Almagro 3

institute: Spanish National Research Center (CNIO)

summary: Despite recent advances in genomic profiling techniques, the precise mechanisms controlling GBM subtypes and their plasticity are not fully unraveled. Here,using transcriptomic data of patient derived stem cell lines we found that FOSL1 is a master regulator of the MES subtype. Depletion of FOSL1 resulted in loss of themesenchymal gene signature (MGS)  in mouse Kras-mutant neural stem cells and in human brain tumor stem cells.

overall_design: Mouse neural stem cells (NSCs) from  GFAP-Tva, LSL_Cas9; Trp53 lox/lox; GFAP-Crep1 mice were transduced with a KrasG12V mutant and then with either Control or Fosl1 gRNAs


title: Single-cell profiling reveals Colonic CD8 Topography in IBD

status: Public on Aug 02 2020

city: Oxford

first: Alison

last: Simmons

address: John Radcliffe Hospital/Headley Way

department: The MRC Weatherall Institute of Molecular Medicine

institute: University of Oxford

summary:
Aims: Single-cell transcriptomic analysis of CD8 T cells in colonic tissue in Health and Ulcerative Colitis
Results: We demonstrated significant heterogeneity within the CD8 population, defined disease associated populations and traced lineage relationships between different T cell subtypes

overall_design: Methods: Colonic biopsies from patients with endoscopically and histologically proven inflamed ulcerative colitis or healthy tissue were taken with informed consent. CD8 T cells were sorted using flow cytometry (FACS) and analysed using the 10X Single-cell RNA sequencing platform to yield matched gene expression, T-Cell receptor (TCR) and selected surface protein (CITE-seq) information across two batches (detailed below).

first: Daniele

last: Corridoni

first: Tarun

last: Gupta

first: Agne

last: Antanaviciute

first: Hubert

last: Slawinki

first: Rory

last: Bowden


title: Constitutive Activation of NF-kB Inducing Kinase (NIK) in the Mesenchymal Lineage Drives Spontaneous Soft Tissue Sarcoma

status: Public on Sep 25 2020

city: Rochester

first: Asha

last: Nair

address: 200 1ST ST SW

institute: Mayo Clinic

summary: Aberrant NF-kB signaling fuels tumor growth in multiple human cancer types including both hematologic and solid malignancies.  Chronic elevated alternative NF-kB signaling can be modeled in transgenic mice upon activation of a conditional NF-kB-inducing kinase (NIK) allele lacking the regulatory TRAF3 binding domain (NT3).  Here, we report that expression of NT3 in the mesenchymal lineage with Osterix (Osx/Sp7)-Cre or Fibroblast Specific Protein (Fsp1)-Cre caused subcutaneous, soft tissue tumors.  These tumors displayed significantly shorter latency and a greater multiple incidence rate in Fsp1-Cre;NT3 compared to Osx-Cre;NT3 mice, regardless of sex.  Histological assessment revealed poorly differentiated solid tumors with some spindled patterns, as well as robust RelB (Relb) immunostaining, confirming activation of alternative NF-kB.  Even though NT3 expression also occurs in the osteolineage in Osx-Cre;NT3 mice, we observed no bony lesions.  The staining profiles and pattern of Cre expression in the two lines pointed to a mesenchymal tumor origin.  Immunohistochemistry revealed that these tumors stain strongly for SMA (Acta2), although vimentin (Vim) staining was uniform only in Osx-Cre;NT3 tumors.  Negative CD45 (Ptprc) and S100 immunostains precluded hematopoietic and melanocytic origins, respectively, while positive staining for CK19 (Krt19), typically associated with epithelia, was found in subpopulations of both tumors.  Principal component, differential expression, and gene ontology analyses revealed that NT3 tumors are distinct from normal mesenchymal tissues and are enriched for NF-kB related biological processes.  We conclude that constitutive activation of the alternative NF-kB pathway in the mesenchymal linage drives spontaneous sarcoma and provides a novel mouse model for NF-kB related sarcomas.

overall_design: Tumors were excised to remove any gross overlaying tissue, and either stored at -20C in RNAlater (R0901; Sigma, USA) or flash frozen in liquid nitrogen, and stored at -80C.  Frozen tumors were pulverized in liquid nitrogen followed by RNA extraction as previously described (PMID:  31246323).  Samples were screened by quantitative real-time PCR (qPCR) for blood- (Hba1, Hba2, and Hbb) and monocyte-specific (CD68) markers to assess low peripheral blood content.  Gene expression was normalized to the levels of Gapdh, a housekeeping gene.  A Nanodrop device (ND-2000; ThermoFisher, USA) and other RNA quality assessments (Bioanalyzer 2100; Agilent Technologies, USA) were used to determine RNA concentration and RNA integrity number (RIN score).  Samples with sufficient RNA yields as well as RIN scores and DV200 scores were prioritized for RNA-Seq analysis as previously described (PMID:  32240779 and 24905804).  Final samples for analysis (n=3 each genotype, all female, Osx-Cre;NT3 = 3 trunk or FSP1-Cre;NT3 = 1 trunk and 2 perineal) had an average RIN score of 7.7 for Osx-Cre;NT3 and 8.5 for FSP1-Cre;NT3.

first: Andre

middle: J

last: van Wijnen

first: Deborah

last: Veis


title: RNA-Seq of Cellulophaga baltica and its phage phi14:2 during the infection process

status: Public on Jul 04 2020

city: Skolkovo

first: Matvei

last: Kolesnik

address: 3 Nobel str

department: Life Sciences

institute: Skoltech

summary: We performed high-througput RNA sequencing of RNA samples extracted from C. baltica cells infected with phi14:2 phage at 40, 90, 140 and 190 minutes after infection. The same experiments were performed with cells treated with host RNA polymerase inhibitor in order to determine which genes rely on viral rifampicin-insensitive RNA polymerase. Using this data, we identified different time classes of phage genes and revealed genes transcribed by phage RNA polymerase.

sample_organism:
Cellulophaga baltica
Cellulophaga phage phi14:2

overall_design: The differences of transcript abundances of phage genes at different stages of infection were analyzed. The ratios of transcript abundances between rifampicin-treated and untreated samples were calculated.

first: Maria

last: Sokolova


title: Transcriptomic Analysis Links the Response of Diverse Hypothalamic Cell Types to Sustained Diabetes Remission Induced by Fibroblast Growth Factor 1 [bulk RNA-seq]

status: Public on Jul 01 2020

city: Copenhagen

first: Tune

middle: Hannes

last: Pers

address: Blegdamsvej 3B

department: Novo Nordisk Foundation Center for Basic Metabolic Research

institute: University of Copenhagen

summary: In rodent models of type 2 diabetes (T2D), sustained remission of hyperglycemia can be induced by a single intracerebroventricular (icv) injection of fibroblast growth factor 1 (FGF1), and the hypothalamus was recently identified as a key target for this effect. To investigate how FGF1 action in this brain area achieves this effect, we combined single-nucleeus RNA sequencing (RNA-seq) with single-cell and traditional RNA-seq to identify >70,000 single-cell transcriptomes from the hypothalamus of diabetic Lepob/ob mice obtained on Days 1, 5 and 42 after icv injection of either FGF1 or vehicle. In addition to robust transcriptomics effects on Agrp neurons, which persisted through Day 42, rapid inhibition of Agrp neurons by FGF1 was shown by both electrophysiology and cFos studies. This effect is predicted to increase hypothalamic melanocortin signaling, and we show that FGF1-induced diabetes remission is melanocortin-dependent, as it was prevented by either genetic or pharmacological blockade of central melanocortin receptors. Combined with dramatic transcriptional and morphological changes induced by icv FGF1 injection in tanycytes, astrocytes and oligodendrocytes, including increased cellular contacts between astrocytes and Agrp neurons, these findings implicate glial-neuron interactions as mediators of the sustained remission of diabetes induced by the hypothalamic action of FGF1 action.

overall_design: bulk RNA-sequencing

first: Marie

middle: A

last: Bentsen

first: Dylan

middle: M

last: Rausch

first: Michael

middle: W

last: Schwartz


title: Enhancer Release and Retargeting” Activates Disease Susceptibility Genes (GRO-Seq)

first: Jiaofang

last: Shao

summary:
Purpose: We used Global run-on sequencing, a gold standard assay to measure transcriptional activity to examine the change of genome transcription in the absence of RAD21 subunit of cohesin, a critical chromatin architectural protein complex. In particularl, we tested how the loss of chromatin looping regulator cohesin would impact the enhancers and their target promoter transcription.
Methods: Transfection of small interfering RNAs (siRNAs) into MCF-7 cells was performed using Lipofectamine 2000 (Life technologies) following manufacturer's instructions. For all experiments, two rounds of siRNA transfection (40nM each time) were performed to achieve higher efficiency. The siRNAs used in this study include: Qiagen Negative Control siRNA (Qiagen Cat# 1027310) and/or Sigma Mission siRNA universal control #2 (SIC002); siRAD21 (Sigma SASI_Hs02_00341219 and SASI_Hs01_00195799). The transfected MCF7 cells were subjected to GRO-seq experiments. GRO-Seq experiments were performed as previously reported (Li et al., 2013, Nature). Briefly, ~10-20 millions of MCF-7 cells were washed 3 times with cold PBS and then sequentially swelled in swelling buffer (10mM Tris-Cl pH7.5, 2mM MgCl2, 3mM CaCl2) for 5 min on ice, harvested, and lysed in lysis buffer (swelling buffer plus 0.5% NP-40 and 10% glycerol). The resultant nuclei were washed one more time with 10mL lysis buffer and finally re-suspended in 100uL of freezing buffer (50mM Tris-Cl pH8.3, 40% glycerol, 5mM MgCl2, 0.1mM EDTA). For the run-on assay, re-suspended nuclei were mixed with an equal volume of reaction buffer (10mM Tris-Cl pH 8.0, 5mM MgCl2, 1mM DTT, 300mM KCl, 20 units of SUPERase-IN, 1% sarkosyl, 500uM ATP, GTP, and Br-UTP, 2uM CTP) and incubated for 5 min at 30°C . The resultant nuclear-run-on RNA (NRO-RNA) was then extracted with TRIzol LS reagent (Life Technologies) following manufacturer’s instructions. NRO-RNA was fragmented to ~300-500nt by alkaline base hydrolysis on ice and followed by treatment with DNase I and Antarctic phosphatase. These fragmented Br-UTP labeled nascent RNA was then immunoprecipitated with an anti-BrdU argarose beads (Sc32323ac, Santa Cruz Biotechnology) in binding buffer (0.5XSSPE, 1mM EDTA, 0.05% tween) for 3 hrs at 4°C with rotation. Subsequently, T4 PNK was used to repair the end of the immunoprecipitated BrU-NRO-RNA, at 37°C for 1hr. The RNA was extracted and precipitated using acidic phenol-chloroform.    cDNA synthesis was performed as per a published method (54) with few modifications. The RNA fragments were subjected to poly-A tailing reaction by poly-A polymerase (NEB) for 30 min at 37°C. Subsequently, reverse transcription was performed using oNTI223 primer and superscript III RT kit (Life Technologies). The cDNA products were separated on a 10% polyacrylamide TBE-urea gel and only those migrated between ~100-500bp were excised and recovered by gel extraction. After that, the first-strand cDNA was circularized by CircLigase (Epicentre) and re-linearized by APE1 (NEB). Re-linearized single strand cDNA (sscDNA) was separated by a 10% polyacrylamide TBE gel as described above and the product of needed size was excised (~170-400bp) for gel extraction. Finally, sscDNA template was amplified by PCR (usually between 10-14 PCR cycles) using the Phusion High-Fidelity enzyme (NEB) according to the manufacturer’s instructions. The resulted library was subjected to deep sequencing.
Results: Using an optimized data analysis workflow, we generated three biological replicates of GRO-seq in the presence or absence of cohesin, and with or without treatment by estrogen.

first: Soohwan

last: Oh

first: Wenbo

last: Li


title: Therapeutic decoupling of MHC-I and PD-L1 expression increases the efficacy of immune checkpoint blockade (SMAC_RNAseq)

city: Boston

first: Shengqing

last: Gu

state: MA

address: 3 Blackfan circle

institute: Dana-Farber Cancer Institute

summary: Immune checkpoint blockade (ICB) therapy has revolutionized the treatment of multiple cancers, but the majority of patients remain refractory due to impaired MHC-I expression. Although the combination of immunotherapy with existing anti-cancer treatments has shown synergy in multiple scenarios, the immunomodulatory effects of conventional therapies remain poorly understood. Here, we integrated CRISPR-mediated genetic screens and data mining of perturbation associated gene expression data to identify drugs that can specifically up-regulate MHC-I without inducing PD-L1. Using FACS-based genome-wide CRISPR screens, we identified Traf3, a critical suppressor of the NF-κB pathway, as a suppressor of MHC-I, but not PD-L1. A Traf3-knockout gene expression signature is associated with better survival in ICB-naive cancer patients and better response to ICB in published clinical trials. From publicly available drug treatment data, we identified SMAC mimetics as the top candidates of having similar transcriptional effects as Traf3-knockout. We experimentally validated that the SMAC mimetic birinapant specifically up-regulated MHC-I, sensitized cancer cells to T-cell-dependent killing, and synergized with ICB in the treatment of established tumors. Our findings provide a strong preclinical rationale for treating MHC-I-low tumors with SMAC mimetics to enhance sensitivity to existing immunotherapy. Moreover, the integrated approach used in this study can be generalized to identify drugs with immunomodulatory effects to enhance immunotherapy efficacy.

overall_design: RNA-seq on tumors treated by birinapant and/or ICB.

first: Wubing

last: Zhang

first: Myles

last: Brown

first: X

last: Liu


title: Therapeutic decoupling of MHC-I and PD-L1 expression increases the efficacy of immune checkpoint blockade

summary: This SuperSeries is composed of the SubSeries listed below.


title: Nfil3, a target of the NACA transcriptional coregulator, affects osteoblast and osteocyte gene expression differentially.

status: Public on Jul 14 2020

city: Montreal

first: Hadla

last: Hariri

state: Quebec

address: 1003 Decarie Boulevard, Shriners Hospital For Children

department: Human Genetics

country: Canada

institute: Shriners Hospital for Children, Canada-Montreal, Department of Human Genetics, McGill University, Montreal,Quebec,Canada

summary: Purpose: Identify PTH-responsive genes in MC3T3-E1 cells. The aim of the study is to identify novel gene targets that are reugulated by PTH signaling in bone, elicting its anabolic effect in osteoblasts. Methods: mRNA expression profiles were compared between Vehicle and PTH(1-34)-treated MC3T3-1 cells, in triplicates, using Illumina HiSeq 2000 PE100 sequncing. qPCR using Taqman probes were used to validate genes of interest. Results: Sequence reads per sample were mapped to the mouse genome (mm10) and differential expression of each gene was determined. Genes that showed differential expression between Vehicle vs PTH(1-34)-treated samples and has a relevant function in bone were further characterized by knockdown strategies in MC3T3-E1 cells. Conclusion: Our study have identified a list of novel genes regulated by PTH siganling. The characterization of their transcription and function in osteoblasts and osteocytes added to our knowledge of PTH signaling in bone and other skeletal tissues.

overall_design: Analysis of total RNA isolated from MC3T3-E1 treated with Vehicle or PTH(1-34).

first: Rene

last: St-Arnaud


title: Therapeutic decoupling of MHC-I and PD-L1 expression increases the efficacy of immune checkpoint blockade (CRISPR_Screen)

overall_design: FACS-based genome-wide CRISPR screens on B16F10 cells for identification of MHC-I and/or PD-L1 regulators


title: Therapeutic decoupling of MHC-I and PD-L1 expression increases the efficacy of immune checkpoint blockade (koTRAF3_RNAseq)

overall_design: RNA-seq on TRAF3-replete or -deficient B16F10 cells that were treated with vehicle or 1ng/ml IFNγ.


title: Therapeutic decoupling of MHC-I and PD-L1 expression increases the efficacy of immune checkpoint blockade (koTRAF3_ATACseq)

overall_design: ATAC-seq on TRAF3-replete or -deficient B16F10 cells that were treated with vehicle or 1ng/ml IFNγ.


title: Transcriptomic Analysis Links the Response of Diverse Hypothalamic Cell Types to Sustained Diabetes Remission Induced by Fibroblast Growth Factor 1

overall_design: Refer to individual Series


title: miR-22 promotes stem cell traits via activating WNT/β-catenin signaling in cSCC

status: Public on Aug 15 2020

city: Tianjin

first: Peitao

address: Qixiangtai Road 22

institute: Tianjin Medical University

summary:
Purpose: Next-generation sequencing (NGS) has revolutionized system-based cell pathway analysis. The purposes of this study were to compare and analyze the expression differences of skin squamous cell line A431 in stem cells after Mir-22 deletion.
Methods: mRNA profiles of  control and case cells were generated by deep sequencing, in duplicate, using Illumina NovaSeq 6000. The sequence reads that passed quality filters were analyzed at the transcript isoform level with two methods:Bowtie2 to map clean reads to reference gene and use HISAT to reference genome.Bowtie2 parameters forSE reads:-q--phred64--sensitive--dpad0--gbar and HISTAparameters forSE reads:-p8--phred64--sensitive-I1-X 1000. qRT–PCR validation was performed using SYBR Green assays
Results: Using an optimized data analysis workflow, we mapped about 24 million sequence reads per sample to the human genome (build hg19) and identified 17000 transcripts in control and case cells with RSEM workflow.  Approximately 5% of the transcripts showed differential expression between  control and case cells, with a fold change ≥1.5 and p value <0.01. Altered expression of 10 candidate genes was confirmed with qRT–PCR, demonstrating the high degree of sensitivity of the RNA-seq method.

first: Shukai

last: Yuan

first: Li

last: Zhao


title: Integrative profiling of early host chromatin accessibility responses in human neutrophils with sensitive pathogen detection [RNA-seq]

status: Public on Jun 30 2020

city: Palo Alto

first: Samuel

last: Yang

state: CA

address: 1201 Welch Road

department: Emergency Medicine

institute: Stanford University School of Medicine

summary: ATAC-seq and RNA-seq of primary human neutrophils challenged with multiple pathogen mimicking ligands and whole organisms.

overall_design: RNA-seq profiles of neutrophils in response to various pathogen ligands and whole organisms

first: Nikhil

last: Ram Mohan

first: Simone

last: Thair


title: Expression data (U133 Plus 2.0) from fibroblast like synoviocytes from patients with rheumatoid arthritis (RA-FLS) stimulated by FasL

city: Kobe

first: Koji

last: Fukuda

address: 7-5-1 Kusunoki, Chuo

department: Department of Orthopaedic Surgery

institute: Kobe University Graduate School of Medicine

summary:
Fas ligand (FasL)/TNFSF6, a member of the tumor necrosis factor (TNF) superfamily, can promote apoptosis in activated primary B cells, T cells, dendritic cells, and synovial fibroblasts through Fas and  is involved in the pathogenesis of autoimmune diseases including rheumatoid arthritis (RA). Meanwhile, decoy receptor 3 (DcR3) competitively binds soluble FasL in addition to TL1A and LIGHT and inhibits the signaling of FasL via Fas. Therefore, FasL-DcR3/Fas signaling may be involved in the pathogenesis of RA.
We hypothesized that FasL regulates the gene expression in RA-FLS. We used to search for genes in which expression in RA-FLS is regulated by FasL.

overall_design: RA-FLS were obtained from 4 RA patients (sample1-4). Each sample was incubated with either 1.0 μg/ml recombinant human FasL protein or phosphate buffered saline diluted with serum-free Opti-MEM medium as non-stimulated control. Gene expression in RA-FLS stimulated by FasL was compared with that of their respective non-stimulated controls.

first: Yasushi

last: Miura

first: Toshihisa

last: Maeda

first: Shinya

last: Hayashi

first: Tomoyuki

last: Matsumoto

first: Ryosuke

last: Kuroda


title: Integrative profiling of early host chromatin accessibility responses in human neutrophils with sensitive pathogen detection


title: Integrative profiling of early host chromatin accessibility responses in human neutrophils with sensitive pathogen detection [ATAC-seq]

overall_design:
ATAC-seq profiles of neutrophils in response to various pathogen ligands and whole organisms

**Please note that the processed *bed files were replaced on March 1, 2021**


title: Chromatin regulates expression of small RNAs to help maintain transposon methylome homeostasis in Arabidopsis

status: Public on Jun 25 2020

city: Vienna

first: Ranjith

last: Papareddy

address: Dr Bohr Gasse 3

country: Austria

institute: Gregor Mendel Institute

summary: We demonstrated that thousands of transposable elements (TEs) produce exceptionally high levels of siRNAs in Arabidopsis thaliana embryos.Depending on whether they are located in euchromatic or heterochromatic regions of the genome, bipartite-classified TEs generate siRNAs throughout embryogenesis according to two distinct patterns. siRNAs are transcribed in embryos and required to direct the re-establishment of DNA methylation on TEs from which they are derived in the new generation.

overall_design:
Examination of  small RNAs and DNA methylation in Arabidopsis embryos and post embryonic tissues

Please note that each processed data generetated from all replicates is linked to the corresponding *Rep1 sample records.

first: Michael

last: Nodine


title: Transcriptomic profiling in NTAPP-exposed human dermal papilla cells.

status: Public on Jun 16 2020

city: Seoul

first: Doyoung

last: Kim

address: 50-1 Yonsei-ro Seodaemun-gu

department: Dermatology

country: South Korea

institute: Yonsei University College of Medicine

summary: To screen biological effects of NTAPP, we performed bulk RNA sequencing on NTAPP-treated human dermal papilla (hDP) cells. Pathway enrichment analysis revealed broad biological effects of NTAPP on hDP cells. Particulaarly, supervised gene ontology analysis suggested that NTAPP-treated hDP cells showed transcriptomic changes in genes associated with Wnt/beta-catenin signaling pathway.

overall_design: Total RNAs were directly purified and isolated from NTAPP-treated hDP cells or non-treated hDP cells.

first: Ji-Hye

last: Hwang

first: Do-Young


title: CircRNA profilings in gastric cancer

status: Public on Jun 12 2020

city: hefei

first: Xiaolin

last: Wang

address: anhui,hefei,huangshan road 443

institute: USTC

summary: To identify GC-related RNAs, we used five GC patients’ fresh tumor tissues and paired adjacent non-tumor tissues for RNA sequencing. Differential expressions of circRNAs and mRNAs were then analyzed.

overall_design: circRNA profiles of 5 paired tumor and paratumor tissues of GC patients.

first: Wang

last: Xiaolin


title: Multiplexed single cell RNA/AbSeq sequencing of ex vivo Treg and ex-Treg from different time points

status: Public on Jun 13 2020

city: Leuven

first: Susan

last: Schlenner

address: Herestraat 49

department: Microbiology, Immunology and Transplantation

country: Belgium

institute: KU Leuven

summary:
Purpose: identify differentially expressed genes between ex-Treg obtained 5, 10 or 15 days post-adoptive transfer into a lymphopenic environment compared to ex vivo Treg in order to identify Treg subset with heightened instability.
Method: Treg from Foxp3YFP-Cre Rosa26RFP (PMID: 18387831, PMID: 17171761) were co-injected with congenically-disparated naive T cells in a 1:1 ratio into Rag1KO mice and ex-Treg were sorted at 15 days, 10 days or 5 days post-transfer. As a control, ex vivo Treg were sorted from Foxp3YFP-Cre Rosa26RFP (d0 sample). Next, samples were stained with multiplexing antibodies using the BD Single-Cell Multiplexing Kit (BD biosciences), then pooled and stained with the following  oligonucleotide-conjugated antibodies: anti‑CD25 (PC61), anti-CD44 (IM7), anti-CD62L (MEL-14), anti-GITR (DTA-1), anti-CD4 (RM4-5), anti‑TCRb (H57-597), anti‑CD69 (H1.2F3), anti‑TIGIT (1G9), anti‑CD95 (Jo2), anti‑CD122 (TM‑1β), anti‑Tim-3 (5D12), anti‑CCR7 (4B12), anti‑CD103 (M290), anti‑CD279 (J43), anti‑CD274 (MIH5), anti‑CD233 (C9B7W), anti‑CD71 (C2), anti‑CD278 (7E7G9), anti‑ITGB7 (M293), anti‑CD137 (1AH2), anti‑CD40 (3/23), anti‑CD3e (145-2C) from BD Biosciences. Single-cell capture and cDNA library preparation was performed using the BD Rhapsody Single-cell analysis system (BD Biosciences), according to the manufacturer’s instructions, using a custom gene panel (591 genes). Sequencing was performed on an Illumina NextSeq500 instrument using a Mid‑Output kit v2.5 (150 cycles, paired-end). Sample demultiplexing, barcode processing, alignment, filtering, UMI counting were done using the standard BD Biosciences Rhapsody analysis pipeline on Seven Bridges (www.sevenbridges.com).
Result: In this study, we showed that ex-Treg downregulate Treg core-specific genes immediatly upon adoptive transfer into a lymphopenic environment. Further, we identified a Treg population enriched for unstable Trg clones.
Conclusion: Our study was able to identify a Treg subset enriched for unstable Treg with plastic potential. This Treg subset appears highly enriched for naïve peripheral-induced Treg.

overall_design: ex vivo Treg and ex-Treg 5, 10 or 15 days post-adoptive transfer into Rag1KO mice were sorted, stained for multiplexing antibodies and stained for Abseq antibodies. Cell capture and single cell RNA sequencing was performed for a custom gene panel using the BD Rhapsody and  Illumina NextSeq500 instruments.

first: Steffie

last: Junius

first: Pierre

last: Lemaitre

first: Adamantios

last: Mavrogiannis

first: Adrian

last: Liston

first: Edyta

last: Kowalczyk


title: mRNA-seq of ARID1B KO and ARID1A/1B DKO livers

status: Public on Jun 10 2020

city: DALLAS

first: Zixi

state: TX

address: 5323 HARRY HINES BLVD

department: CRI

institute: UT SOUTHWESTERN MEDICAL CENTER

summary: ARID1B KO in liver leads to minimal gene expression changes, whereas ARID1A/1B DKO leads to global gene expression changes.

overall_design: mRNA-seq of liver samples.

first: Kenian

last: Chen

first: Hao

last: Zhu


title: Translational adaptation to heat stress is mediated by 5-methylcytosine RNA modification in Caenorhabditis elegans

city: cambridge

first: Jonathan

middle: Lewis

last: Price

address: The Sanger Building Tennis Court Road

department: Biochemistry

country: United Kingdom

institute: University of Cambridge

summary: In order to fulfil a wide range of cellular functions, RNA molecules contain, in addition to the four canonical bases, a large number of chemically modified structures. Among those, the methylation of carbon-5 of cytosines (m5C) is an abundant, widespread mark in different classes of RNAs. m5C is catalysed by DNMT2 and by seven members of the NSUN family of enzymes. While some m5C-methyltransferases have been studied individually and implicated in phenotypes observed in organisms ranging from yeast to humans, it remains unclear how the m5C methylome acts to sustain development in homeostasis and stress. Here, using Caenorhabditis elegans as a model organism, we developed the first organism devoid of detectable m5C in RNA, providing strong evidence that this modification and its derivatives are non-essential for the nematode’s development and fertility under standard laboratory conditions. We further used this genetic tool to determine the localisation of m5C sites in different RNA classes, and showed that NSUN-4 acts as a multisite-specific tRNA/rRNA methyltransferase in the mitochondria. In addition, we show that m5C is required for adaptation to temperature changes, as animals devoid of this modification present temperature-sensitive fertility defects. At the molecular level, we show that loss of m5C leads to increased ribosome pausing at triplets encoding for leucine and proline, the most frequently methylated tRNA isoacceptors in C. elegans. This is correlated with reduced translation efficiency of transcripts enriched in these amino acids. Finally, we found translation of Leu-UUG codons to be the most strongly affected upon heat shock, suggesting a role of m5C wobble methylation in the adaptation to heat stress.

overall_design: RNASeq and Ribosome profilling from 4 samples of C. elegans. WT and NSUN mutants from 20 and 27 degrees celcius.

first: Isabela

last: Navarro

first: Francesca

last: Tuorto

first: David

last: Jordan

first: Carine

last: Legrand

first: Fabian

last: Braukmann

first: Alan

last: Hendrick

first: Alper

last: Akay

first: Annika

last: Kotter

first: Mark

last: Helm

first: Frank

last: Lyko

first: Eric

last: Miska


title: Large-scale evaluation of renewable PRCP monoclonal antibodies by ChIP-exo, STORM, and …

status: Public on Jun 09 2020

city: Durham

first: Bryan

last: Venters

state: NC

address: 6 Davis Drive

country: USA

institute: EpiCypher

summary: The study of chromatin and its organization involves use of antibodies directed against thousands of distinct proteins spanning numerous isoforms and post-translationally modified variants in the human nucleus. Antibodies are essential where epitope-tagging is impractical (e.g., human tissue, cell lines from a wide variety of origins). However, antibody reliability is problematic. The Protein Capture Reagents Program (PCRP) has generated about a thousand renewable monoclonal antibodies that are expected to be more reliable, but these reagents have not been field tested.  Since they were generated primarily against chromatin proteins, we tested them in a variety of chromatin-based assays. 887 unique antibodies against 681 unique chromatin targets were assayed by ChIP-exo. A subset were further tested in ChIP-seq, CUT&RUN, STORM super-resolution microscopy, immunoblots, and PBM protein-DNA interaction assays. While this represents only a small portion of the antibody validation/utility space, and noting that optimization may be needed in each assay, our findings suggest that 10-20% of the PCRP-derived antibodies have utility in a particular biochemical assay. We also describe validation strategies.

overall_design: We performed CUT&RUN usking K562 extracted nuclei on 40 TFs using 40 PCRP sourced antibodies

first: Katherine

last: Novitzky

first: Eileen

last: McAnarney

first: Michael-C.

last: Keogh

first: William

middle: K

last: Lai

first: Thomas

last: Blanda

first: Kylie

last: Bocklund

first: Prashant

last: Kuntala

first: Joshua

last: Mairose

first: Sarah

last: Dweikat

first: Katelyn

last: Mistretta

first: B

last: Pugh

first: Avani

middle: P

last: Shaw

first: Marta

last: Iwanaszko

first: Edwin

last: Smith

first: Ali

last: Shilatifard

first: Luca

last: Mariani

first: James

middle: T

last: Anderson

first: Sabrina

last: Phanor

first: Martha

middle: L

last: Bulyk

first: Gerson

last: Rothschild

first: Sebastian

last: Fernando

first: Wanwei

last: Zhang

first: Uttiya

last: Basu